Amino Acid Substitution Rates Research Articles

Long-term persistence of infection in chimpanzees inoculated with an infectious hepatitis C virus clone is associated with a decrease in the viral amino acid substitution rate and low levels of heterogeneity.

Two chimpanzees, 1535 and 1536, became persistently infected following inoculation with RNA transcripts from cDNA clones of hepatitis C virus (HCV). Analysis of the HCV genomes from both animals showed an accumulation of amino acid substitutions over time. The appearance of substitutions in the envelope genes was associated with increased antienvelope antibody titers. However, extensive mutations were not incorporated into hypervariable region 1 (HVR1). A comparison of the nonsynonymous substitution rate/synonymous substitution rate was made at various time points to analyze selective pressure. The highest level of selective pressure occurred during the acute phase and decreased as the infection continued. The nonsynonymous substitution rate was initially higher than the synonymous substitution rate but decreased over time from 3.3 x 10(-3) (chimpanzee 1535) and 3.2 x 10(-3) (chimpanzee 1536) substitutions/site/year at week 26 to 1.4 x 10(-3) (chimpanzee 1535) and 1.7 x 10(-3) (chimpanzee 1536) at week 216, while the synonymous substitution rate remained steady at approximately 1 x 10(-3) substitutions/site/year. Analysis of PCR products using single-stranded conformational polymorphism indicated a low level of heterogeneity in the viral genome. The results of these studies confirm that the persistence of infection is not solely due to changes in HVR1 or heterogeneity and that the majority of variants observed in natural infections could not arise simply through mutation during the time period most humans and chimpanzees are observed. These data also indicate that immune pressure and selection continue throughout the chronic phase.

Analysis of mitochondrial DNA protein-coding region in the Yeso Sika deer (Cervus nippon yesoensis)

In the present study, mitochondrial DNA sequences of the Yeso Sika deer (Cervus nippon yesoensis) were studied. Specifically, protein-coding genes as mitochondrial NADH dehydrogenase subunits (ND1, ND2, ND3, ND4L, ND4, ND5 and ND6), cytochrome c oxidase subunits (CO I and CO III), ATP synthase subunits (ATPase8 and ATPase6) and cytochrome b. Also, phylogenetic analyses on eight mammalian species were performed, including the Muntjac deer (Muntiacus reevesi). The rate of amino-acid substitution was lowest (3.74%) between Yeso Sika deer and Muntjac deer, and the values between Yeso Sika deer and other species (sheep, cattle, horse, pig, mouse, human and chimpanzee) were 6.63%, 7.30%, 12.55%, 13.03%, 23.59%, 24.82% and 25.04%, respectively. Among them, the highest value of divergence was recognized in ATPase8, and the second structure of ATPase8 showed a difference between the Yeso Sika deer and Muntjac deer as a result of the substitution of 34HisTyr and 49ThrIle. In addition, we identified a substitution of an amino-acid sequence (19ThrAla) between the Yeso Sika deer and Yakushima Sika deer (C. n. yakushimae). From these results, ATPase8 was also a variable region in Cervidae.

Rapid evolution of mammalian X-linked testis-expressed homeobox genes.

Homeobox genes encode transcription factors that function in various developmental processes and are usually evolutionarily conserved in their sequences. However, two X-chromosome-linked testis-expressed homeobox genes, one from rodents and the other from fruit flies, are known to evolve rapidly under positive Darwinian selection. Here we report yet another case, from primates. TGIFLX is an X-linked homeobox gene that originated by retroposition of the autosomal gene TGIF2, most likely in a common ancestor of rodents and primates. While TGIF2 is ubiquitously expressed, TGIFLX is exclusively expressed in adult testis. A comparison of the TGIFLX sequences among 16 anthropoid primates revealed a significantly higher rate of nonsynonymous nucleotide substitution (d(N)) than synonymous substitution (d(S)), strongly suggesting the action of positive selection. Although the high d(N)/d(S) ratio is most evident outside the homeobox, the homeobox has a d(N)/d(S) of approximately 0.89 and includes two codons that are likely under selection. Furthermore, the rate of radical amino acid substitutions that alter amino acid charge is significantly greater than that of conservative substitutions, suggesting that the selection promotes diversity of the protein charge profile. More interestingly, an analysis of 64 orthologous homeobox genes from humans and mice shows substantially higher rates of amino acid substitution in X-linked testis-expressed genes than in other genes. These results suggest a general pattern of rapid evolution of mammalian X-linked testis-expressed homeobox genes. Although the physiological function of and the exact selective agent on TGIFLX and other rapidly evolving homeobox genes are unclear, the common expression pattern of these transcription factor genes led us to conjecture that the selection is related to one or more aspects of male reproduction and may contribute to speciation.

Molecular evolution of the SARS coronavirus during the course of the SARS epidemic in China.

Sixty-one SARS coronavirus genomic sequences derived from the early, middle, and late phases of the severe acute respiratory syndrome (SARS) epidemic were analyzed together with two viral sequences from palm civets. Genotypes characteristic of each phase were discovered, and the earliest genotypes were similar to the animal SARS-like coronaviruses. Major deletions were observed in the Orf8 region of the genome, both at the start and the end of the epidemic. The neutral mutation rate of the viral genome was constant but the amino acid substitution rate of the coding sequences slowed during the course of the epidemic. The spike protein showed the strongest initial responses to positive selection pressures, followed by subsequent purifying selection and eventual stabilization.

Serological pattern ‘anti-hepatitis B core alone’ in HIV or hepatitis C virus-infected patients is not fully explained by hepatitis B surface antigen mutants

We determined the frequency of hepatitis B virus (HBV) occult infection and studied S gene mutations in HIV or hepatitis C virus-co-infected patients with persistent pattern 'anti-hepatitis B core alone'. HBV DNA was particularly frequent in HIV-positive patients. Sequencing of the S gene showed that only two HIV-infected patients harboured mutants in the major hydrophilic loop; the absence of hepatitis B surface antigen detection could not be explained by diagnostic escape mutants.

Molecular evolution of the phytochrome gene family in sorghum: changing rates of synonymous and replacement evolution.

The photoreceptor phytochromes, encoded by a small gene family, are responsible for controlling the expression of a number of light-responsive genes and photomorphogenic events, including agronomically important phenotypes such as flowering time and shade-avoidance behavior. The understanding and control of flowering time are particularly important goals in sorghum cultivar development for diverse environments, and naturally occurring variation in the phytochrome genes might prove useful in breeding programs. Also of interest is whether variation observed at the phytochrome loci in domesticated sorghum, or in particular races, is a result of human selection. Population genetic studies can reveal evidence of such selection in patterns of polymorphism and divergence. In this study we report a population genetic analysis of the PHY gene family in Sorghum bicolor (L.) Moench in a diverse panel including both cultivated and wild accessions. We show that the level of nucleotide variation in all gene family members is about half the average for this species, consistent with purifying selection acting on these loci. However, the rate of amino acid substitution is accelerated at PHYC compared to the other two loci. In comparisons to a closely related sorghum species, PHYC shows a pattern of intermediate frequency amino acid changes that differ from the patterns observed in comparisons across longer evolutionary distances. There is also a departure from expected patterns of polymorphism and divergence at synonymous sites in PHYC, although the data do not fit a simple model of directional or diversifying selection. Cultivated sorghum has a level of variation similar to that of wild relatives (ssp. verticilliflorum), but many polymorphisms are subspecies-specific, including several amino acid variants.

Molecular clock in neutral protein evolution

BackgroundA frequent observation in molecular evolution is that amino-acid substitution rates show an index of dispersion (that is, ratio of variance to mean) substantially larger than one. This observation has been termed the overdispersed molecular clock. On the basis of in silico protein-evolution experiments, Bastolla and coworkers recently proposed an explanation for this observation: Proteins drift in neutral space, and can temporarily get trapped in regions of substantially reduced neutrality. In these regions, substitution rates are suppressed, which results in an overall substitution process that is not Poissonian. However, the simulation method of Bastolla et al. is representative only for cases in which the product of mutation rate μ and population size Ne is small. How the substitution process behaves when μNe is large is not known.ResultsHere, I study the behavior of the molecular clock in in silico protein evolution as a function of mutation rate and population size. I find that the index of dispersion decays with increasing μNe, and approaches 1 for large μNe . This observation can be explained with the selective pressure for mutational robustness, which is effective when μNe is large. This pressure keeps the population out of low-neutrality traps, and thus steadies the ticking of the molecular clock.ConclusionsThe molecular clock in neutral protein evolution can fall into two distinct regimes, a strongly overdispersed one for small μNe, and a mostly Poissonian one for large μNe. The former is relevant for the majority of organisms in the plant and animal kingdom, and the latter may be relevant for RNA viruses.

Phylogenetic estimation of context-dependent substitution rates by maximum likelihood.

Nucleotide substitution in both coding and noncoding regions is context-dependent, in the sense that substitution rates depend on the identity of neighboring bases. Context-dependent substitution has been modeled in the case of two sequences and an unrooted phylogenetic tree, but it has only been accommodated in limited ways with more general phylogenies. In this article, extensions are presented to standard phylogenetic models that allow for better handling of context-dependent substitution, yet still permit exact inference at reasonable computational cost. The new models improve goodness of fit substantially for both coding and noncoding data. Considering context dependence leads to much larger improvements than does using a richer substitution model or allowing for rate variation across sites, under the assumption of site independence. The observed improvements appear to derive from three separate properties of the models: their explicit characterization of context-dependent substitution within N-tuples of adjacent sites, their ability to accommodate overlapping N-tuples, and their rich parameterization of the substitution process. Parameter estimation is accomplished using an expectation maximization algorithm, with a quasi-Newton algorithm for the maximization step; this approach is shown to be preferable to ordinary Newton methods for parameter-rich models. Overlapping tuples are efficiently handled by assuming Markov dependence of the observed bases at each site on those at the N - 1 preceding sites, and the required conditional probabilities are computed with an extension of Felsenstein's algorithm. Estimated substitution rates based on a data set of about 160,000 noncoding sites in mammalian genomes indicate a pronounced CpG effect, but they also suggest a complex overall pattern of context-dependent substitution, comprising a variety of subtle effects. Estimates based on about 3 million sites in coding regions demonstrate that amino acid substitution rates can be learned at the nucleotide level, and suggest that context effects across codon boundaries are significant.

Patterns of intra- and interhost nonsynonymous variation reveal strong purifying selection in dengue virus.

Considerable uncertainty surrounds the evolutionary rates of and selection pressures acting on arthropod-borne RNA viruses (arboviruses). In particular, it is unclear why arboviruses such as dengue virus show substantial genetic variation within individual humans and mosquitoes yet low long-term rates of amino acid substitution. To address this question, I compared patterns of nonsynonymous variation in populations of dengue virus sampled at different levels of evolutionary divergence. Although nonsynonymous variation was abundant in viral populations within individual humans, there was a marked reduction in the frequency of nonsynonymous mutations in interhost comparisons. Moreover, intrahost genetic variation corresponded to a random pattern of mutation, and most of the sites that exhibited nonsynonymous variation within hosts were invariant at deeper phylogenetic levels. This loss of long-term nonsynonymous variation is the signature of extensive purifying selection such that more than 90% of all nonsynonymous mutations are deleterious. Consequently, although arboviruses are able to successfully adapt to diverse cell types, they are characterized by a high rate of deleterious mutation.

Using protein design for homology detection and active site searches.

We describe a method of designing artificial sequences that resemble naturally occurring sequences in terms of their compatibility with a template structure and its functional constraints. The design procedure is a Monte Carlo simulation of amino acid substitution process. The selective fixation of substitutions is dictated by a simple scoring function derived from the template structure and a multiple alignment of its homologs. Designed sequences represent an enlargement of sequence space around native sequences. We show that the use of designed sequences improves the performance of profile-based homology detection. The difference in position-specific conservation between designed sequences and native sequences is helpful for prediction of functionally important residues. Our sequence selection criteria in evolutionary simulations introduce amino acid substitution rate variation among sites in a natural way, providing a better model to test phylogenetic methods.

Identification and characterization of mouse Erbb2 gene in silico

The PPP1R1B-STARD3-TCAP-PNMT-MGC9753-ERBB2-MGC14832-GRB7 locus on human chromosome 17q12 is frequently amplified in human gastric and breast cancer. We have recently identified and characterized human MGC9753 (also known as wild-type CAB2) and mouse Mgc9753. Here, we identified and characterized mouse Erbb2 gene by using bioinformatics. BLAST programs revealed that mouse AK031099 cDNA was derived from mouse Erbb2 gene. Because AK031099 cDNA showed 806 C-->A nucleotide substitution compared with mouse genome draft sequences and mouse Erbb2 ESTs, the nucleotide sequence of mouse Erbb2 cDNA was determined in silico by correcting 806 A of AK031099 cDNA to C. Nucleotide position 48-3818 of mouse Erbb2 cDNA was the coding region. Mouse Erbb2 gene, consisting of 27 exons, was located within the Ppp1r1b-Grb7 locus on the mouse chromosome 11. Mouse Erbb2 protein (1256 aa) showed 87.5% total-amino-acid identity with human ERBB2 protein, and 95.2% total-amino-acid identity with rat Erbb2 protein. Mouse Ppp1r1b-Grb7 locus and human Ppp1r1b-Grb7 locus were evolutionarily conserved in the order and the orientation of genes therein. Nucleotide and amino-acid substitution rates of Neurod2 located centromeric to the Ppp1r1b-Grb7 locus were significantly lower than others within the Ppp1r1b-Grb7 locus. This is the first report on the complete coding sequence of mouse Erbb2 gene as well as on the comprehensive comparison of Ppp1r1b-Grb7 locus within the human and mouse genomes.

Adaptive evolution of the water stress-induced gene Asr2 in Lycopersicon species dwelling in arid habitats.

The Asr2 gene encodes a putative transcription factor that is up-regulated in leaves and roots of tomato plants exposed to water-deficit stress. This gene was first cloned and characterized in a cultivar of commercial tomato (Lycopersicon esculentum cv. Ailsa Craig). In this work, we report the complete coding sequences of the orthologous Asr2 genes in six wild tomato lineages: L. hirsutum, L. cheesmanii, L. esculentum v. cerasiforme, L. chilense, L. peruvianum v. humifusum and L. peruvianum f. glandulosum. Estimates of the Ka/Ks ratio (omega) in pairwise comparisons within the genus Lycopersicon were equal or greater than 1 (a signature of adaptive evolution) when involving L. chilense and L. peruvianum v. humifusum. Interestingly, these two species are distinct from the others in their adaptation to dry habitats. We also mapped the detected substitutions onto a phylogenetic tree of the genus Lycopersicon. Remarkably, there are two and three amino acid substitutions, which contrast with the absence of synonymous substitutions along the terminal branches leading to L. chilense and L. peruvianum v. humifusum, respectively. Likelihood ratio tests confirmed that omega values in the branches leading to these species are significantly different from the remaining branches of the tree. Moreover, inferred changes in the branches leading to these species that inhabit dry areas are nonconservative and may be associated with dramatic alterations in ASR2 protein conformation. In this work, we demonstrate accelerated rates of amino acid substitutions in the Asr2 gene of tomato lineages living in dry habitats, thus giving support to the hypothesis of adaptive Darwinian evolution.

Speciation and inversions: chimps and humans.

A new set of models has resurrected a role for chromosomal inversions in the formation of new species. Traditional models, which are generally considered to be unlikely in most cases, had imagined that inversions might aid speciation by directly causing low hybrid fitness. In contrast, the newer models focus on the effect that inversions have on local recombination rates. A test of these models found a strikingly high rate of amino-acid substitution within regions where humans and chimpanzees differ by inversions, suggesting perhaps that our ancestral species underwent a divergence process in which gene flow and inversions played a key role. However, it remains uncertain whether this interesting finding is actually consistent with the proposed model.

Identification and characterization of mouse Ppfia1 gene in silico

Array CGH combined with mRNA microarray analyses was successfully applied for genome-wide screening of proto-oncogenes and tumor suppressor genes in 2002. The CCND1-ORAOV1-FGF19-FGF4-FGF3-FLJ10261-FADD-PPFIA1-EMS1 locus on human chromosome 11q13 is one of the most frequently amplified regions within the human genome. Here, we identified and characterized mouse Ppfia1 gene by using bioinformatics. Nucleotide sequence of mouse Ppfia1 cDNA was determined in silico by assembling nucleotide sequences of ESTs BY727670, CA327608, BU708520, BQ886535, and a 5'-truncated partial cDNA BC038349. Mouse Ppfia1 gene, consisting of 28 exons, was located between Fadd and Ems1 (also known as Cttn) genes on mouse chromosome 7. Mouse Ppfia1 (1201 aa) and human PPFIA1 (1202 aa), showing 95.8% total-amino-acid identity, were found to consist of MAH (myosin heavy chain tail and ATPase homologous) domain and three SAM (sterile alpha motif) domains. MAH domain is implicated in the homo- or hetero-oligomer formation through the coiled-coil interaction, while SAM domain is implicated in the interaction with other proteins. Mouse Ccnd1-Ems1 locus and human CCND1-EMS1 locus were evolutionarily conserved in the order and the orientation of genes therein. Nucleotide and amino-acid substitution rates of Ccnd1, Ppfia1 and Ems1 genes located near both ends of the Ccnd1-Ems1 locus were relatively lower than those in the middle part of the locus. This is the first report on mouse Ppfia1 gene as well as comprehensive comparison of CCND1-EMS1 locus within the human and mouse genomes.

Adaptation shapes patterns of genome evolution on sexual and asexual chromosomes in Drosophila.

What advantage might sexual recombination confer? Population genetics theory predicts that asexual genomes are less efficient at eliminating deleterious mutations and incorporating beneficial alleles. Here, I compare patterns of genome evolution in a 40-kb gene-rich region on homologous neo-sex chromosomes of Drosophila miranda. Genes on the non-recombining neo-Y show various signs of degeneration, including transposable-element insertions, frameshift mutations and a higher rate of amino-acid substitution. In contrast, loci on the recombining neo-X show intact open reading frames and generally low rates of amino-acid substitution. One exceptional gene on the neo-X shows evidence for adaptive protein evolution, affecting patterns of variability at neighboring regions along the chromosome. These findings illustrate the limits to natural selection in an asexual genome. Deleterious mutations, including repetitive DNA, accumulate on a non-recombining chromosome, whereas rapid protein evolution due to positive selection is confined to the recombining homolog.

Recombination cluster around FGFR2-WDR11-HTPAPL locus on human chromosome 10q26

FGFR2 is an oncogene amplified in diffuse-type gastric cancer, and WDR11 is a tumor suppressor gene disrupted in glial tumor. WDR11-FGFR2 locus on human chromosome 10q26 is one of cancer-related recombination hot spots. In this study, we investigated recombination and nucleotide substitution around the WDR11-FGFR2 locus during evolution by using bioinformatics. Inter-chromosomal comparison revealed that the human BAG3-FGFR2-TACC2 region was paralogous to the human BAG4-FGFR1-TACC1 region. Inter-specific comparison on the BAG3-FGFR2-TACC2 region revealed that HTPAPL-WDR11-FGFR2 locus containing species-specific insertion or deletion was one of evolutionary recombination hot spots. Between human and mouse, coding-region nucleotide substitution rate and amino-acid substitution rate were significantly lower in the HTPAPL-WDR11-FGFR2 locus than in the surrounding locus (P<0.0001). The HTPAPL-WDR11-FGFR2 locus was more susceptible to recombination than to nucleotide substitution. Detailed comparison of human and mouse genomes could identify evolutionary recombination hot spots overlooked during gross comparison of human and mouse genomes. Because DNA double-strand break is the initial step in various types of recombination including chromosomal translocation, rearrangement, deletion, gene amplification, retroviral integration and retrotransposition, it is reasonable that the HTPAPL-WDR11-FGFR2 locus is the recombination hot spot during evolution as well as during carcinogenesis. Therefore, comparative genomics might be applicable to identification of recombination hot spots and genes related to cancer.

Protein evolution: structure‐function relationships of the oncogene beta‐catenin in the evolution of multicellular animals

Beta-catenin functions as a cytoskeletal linker protein in cadherin-mediated adhesion and as a signal mediator in wnt-signal transduction pathways. We use a novel integrative approach, combining evolutionary, genomic, and three-dimensional structural data to analyze and trace the structural and functional evolution of beta-catenin genes. This approach also enabled us to examine the effects of gene duplication on the structure and function of beta-catenin genes in Drosophila, C. elegans, and vertebrates. By sampling a large number of different taxa, we identified both ancestral and derived motifs and residues within the different regions of the beta-catenin proteins. Projecting amino acid substitutions onto the three- dimensional structure established for mouse beta-catenin, we identified specific domains that exhibit loss and gain of selective constraints during beta catenin evolution. Structural changes, changes in the amino acid substitution rate, and the appearance of novel functional domains in beta-catenin can be mapped to specific branches on the metazoan tree. Together, our analyses suggest that a single, beta-catenin gene fulfilled both adhesion and signaling functions in the last common ancestor of metazoans some 700 million years ago. In addition, gene duplications facilitated the evolution of beta-catenins with novel functions and allowed the evolution of multiple, single-function proteins (cell adhesion or wnt-signaling) from the ancestral, dual-function protein. Integrative methods such as those we have applied here, utilizing the 'natural experiments' present in animal diversity, can be employed to identify novel and shared functional motifs and residues in virtually any protein among the proteomes of model systems and humans.

Partitioning the variation in mammalian substitution rates.

We have used analysis of variance to partition the variation in synonymous and amino acid substitution rates between three effects (gene, lineage, and a gene-by-lineage interaction) in mammalian nuclear and mitochondrial genes. We find that gene effects are stronger for amino acid substitution rates than for synonymous substitution rates and that lineage effects are stronger for synonymous substitution rates than for amino acid substitution rates. Gene-by-lineage interactions, equivalent to overdispersion corrected for lineage effects, are found in amino acid substitutions but not in synonymous substitutions. The variance in the ratio of amino acid and synonymous substitution rates is dominated by gene effects, but there is also a significant gene-by-lineage interaction.

Different evolutionary patterns between young duplicate genes in the human genome.

Following gene duplication, two duplicate genes may experience relaxed functional constraints or acquire different mutations, and may also diverge in function. Whether the two copies will evolve in different patterns remains unclear, however, because previous studies have reached conflicting conclusions. In order to resolve this issue, by providing a general picture, we studied 250 independent pairs of young duplicate genes from the whole human genome. We showed that nearly 60% of the young duplicate gene pairs have evolved at the amino-acid level at significantly different rates from each other. More than 25% of these gene pairs also showed significantly different ratios of nonsynonymous to synonymous rates (Ka/Ks ratios). Moreover, duplicate pairs with different rates of amino-acid substitution also tend to differ in the Ka/Ks ratio, with the fast-evolving copy tending to have a slightly higher Ks than the slow-evolving one. Lastly, a substantial portion of fast-evolving copies have accumulated amino-acid substitutions evenly across the protein sequences, whereas most of the slow-evolving copies exhibit uneven substitution patterns. Our results suggest that duplicate genes tend to evolve in different patterns following the duplication event. One copy evolves faster than the other and accumulates amino-acid substitutions evenly across the sequence, whereas the other copy evolves more slowly and accumulates amino-acid substitutions unevenly across the sequence. Such different evolutionary patterns may be largely due to different functional constraints on the two copies.

Acceleration of genomic evolution caused by enhanced mutation rate in endocellular symbionts.

Endosymbionts, which are widely observed in nature, have undergone reductive genome evolution because of their long-term intracellular lifestyle. Here we compared the complete genome sequences of two different endosymbionts, Buchnera and a protist mitochondrion, with their close relatives to study the evolutionary rates of functional genes in endosymbionts. The results indicate that the rate of amino acid substitution is two times higher in symbionts than in their relatives. This rate increase was observed uniformly among different functional classes of genes, although strong purifying selection may have counterbalanced the rate increase in a few cases. Our data suggest that, contrary to current views, neither the Muller's ratchet effect nor the slightly deleterious mutation theory sufficiently accounts for the elevated evolutionary rate. Rather, the elevated evolutionary rate appears to be mainly due to enhanced mutation rate, although the possibility of relaxation of purifying selection cannot be ruled out.